Bacterial pathogens elaborate a diverse array of toxins that disrupt the homeostasis of host tissues. Toxins can directly damage host cells, but several enhance virulence by dysregulating the host response and causing inflammatory tissue injury. Furthermore, in many infectious diseases, accumulating evidence suggests that colonization of pathogenic organisms instigates disease, but disease progression is largely mediated by the dysregulated host response to infection. The central goal of this R21 proposal is to explore how heparan sulfate proteoglycans (HSPGs) modulate the host response to Staphylococcus aureus ?-toxin, an exotoxin virulence factor implicated in many staphylococcal diseases. We found in preliminary studies that syndecan-1 null (Sdc1-/-) mice show significantly increased inflammatory lung injury when challenged intranasally (i.n.) with S. aureus ?-toxin. Syndecan-1 is the major cell surface HSPG of type II epithelial cells in the lung, and ?-toxin is an established virulence factor for S. aureus pneumonia. Importantly, the lung injury phenotypes were rescued by i.n. administration of purified syndecan-1 ectodomains, heparan sulfate (HS) or sulfated HS fragments, but not by chondroitin sulfate (CS), core protein devoid of HS and CS chains or low sulfated HS fragments, indicating that sulfated domains in syndecan-1 HS protect against ?-toxin-induced lung injury. However, the critical structural features that enable syndecan-1 HS to function in this manner and the biological target of syndecan-1 in ?-toxin-induced lung injury are unknown. This project will fill these mechanistic gaps by precisely defining the HS sulfation code that enables Sdc1 to prominently and specifically inhibit ?-toxin-induced lung injury (aim 1), and by exploring the role of Sdc1 interactions with innate immune cells and cytokines in the dysregulated host response to ?-toxin. The culminating goal is to provide a molecular, glycobiological, and cellular basis for HSPGs in toxin-induced lung injury for the translational development of new diagnostics and therapeutics for bacterial pneumonia.